This invention relates to a new use of an immunosuppressant. More specifically, this invention relates to a new use of immunosuppressant as matrix metallproteinases production inhibitor (hereinafter, referred to as MMP-production inhibitor).
Matrix metalloproteinases (hereinafter, referred to as MMPs) are a large family of Zn2+ endopeptidases that include 72 and 92 kDa gelatinase, collagenase, stromelysin and membrane-bound MMPs. They are expressed in inflammatory coditions and collectively capable of degrading most connective tissues. MMPs, such as gelatinase (MMP-2, MMP-9), stromelysin (MMP-3) and collagenase (MMP-1, MMP-8, MMP-13), are involved in tissue matrix degradation and have been implicated in many pathological conditions involving abnormal connective tissue and basement membrane matrix metabolism.
This invention provides a new use of an immunosuppressant as MMP-production inhibitor.
Further, this invention provide a new MMP-production inhibitor comprising an immunosuppressant as an active ingredient.
Still further, this invention provide a new use of an immunosuppressant for manufacturing a medicament for preventing or treating MMP-mediated diseases, and a new method by using its medicament and a medicament therefor.
Various immunosuppressants have already been known. For example, it is well known that cyclosporins and tacrolimus (FK506), and their derivatives, possess a strong immunosuppressive activity, which were shown in, for example, J. Antibiotics 40(1987), 1256-1265, U.S. Pat. No. 4,929,611, and so on.
The inventors of this invention have surprisingly found that the immunosuppressant mentioned herein below has a new activity, i.e., MMP-production inhibitory activity.
The xe2x80x9cimmunosuppressantxe2x80x9d used in the present invention should not be limited.
One example of the immunosuppressant is macrolides of the following formula (I). 
(wherein each of adjacent pairs of R1 and R2, R3 and R4, and R5 and R6 independently
(a) is two adjacent hydrogen atoms, but R2 may also be an alkyl group or
(b) may form another bond formed between the carbon atoms to which they are attached;
R7 is a hydrogen atom, a hydroxy group, a protected hydroxy group, or an alkoxy group, or an oxo group together with R1;
R8 and R9 are independently a hydrogen atom or a hydroxy group;
R10 is a hydrogen atom, an alkyl group, an alkyl group substituted by one or more hydroxy groups, an alkenyl group, an alkenyl group substituted by one or more hydroxy groups, or an alkyl group substituted by an oxo group;
X is an oxo group, (a hydrogen atom and a hydroxy group), (a hydrogen atom and a hydrogen atom), or a group represented by the formula xe2x80x94CH2Oxe2x80x94;
Y is an oxo group, (a hydrogen atom and a hydroxy group), (a hydrogen atom and a hydrogen atom), or a group represented by the formula Nxe2x80x94NR11R12 or Nxe2x80x94OR13;
R11 and R12 are independently a hydrogen atom, an alkyl group, an aryl group or a tosyl group;
R13, R14, R15, R16, R17, R18, R19, R22 and R23 are independently a hydrogen atom or an alkyl group;
R24 is an optionally substituted ring system which may contain one or more heteroatoms;
n is an integer of 1 or 2; and
in addition to the above definitions, Y, R10 and R23, together with the carbon atoms to which they are attached, may represent a saturated or unsaturated 5- or 6-membered nitrogen, sulfur and/or oxygen containing heterocyclic ring optionally substituted by one or more groups selected from the group consisting of an alkyl, a hydroxy, an alkoxy, a benzyl, a group of the formula xe2x80x94CH2Se(C6H5), and an alkyl substituted by one or more hydroxy groups.
Preferable R24 may be cyclo(C5-7)alkyl group, and the following ones can be exemplified.
(a) a 3,4-di-oxo-cyclohexyl group;
(b) a 3-R20-4-R21-cyclohexyl group,
in which R20 is hydroxy, an alkoxy group, an oxo group, or a xe2x80x94OCH2OCH2CH2OCH3 group, and
R21 is hydroxy, xe2x80x94OCN, an alkoxy group, a heteroaryloxy which may be substituted by suitable substituents, a xe2x80x94OCH2OCH2CH2OCH3 group, a protected hydroxy group, chloro, bromo, iodo, aminooxalyloxy, an azido group, p-tolyloxythiocarbonyloxy, or R25R26CHCOOxe2x80x94,
xe2x80x83in which R25 is optionally protected hydroxy or protected amino, and
R26 is hydrogen or methyl, or
xe2x80x83R20 and R21 together form an oxygen atom in an epoxide ring; or
(c) cyclopentyl group substituted by methoxymethyl, optionally protected hydroxymethyl, acyloxymethyl
(in which the acyl moiety optionally contains either a dimethylamino group which may be quaternized, or a carboxy group which may be esterified), one or more amino and/or hydroxy groups which may be protected, or aminooxalyloxymethyl. A preferred example is a 2-formyl-cyclopentyl group.
The definitions used in the above general formula (I) and the specific and preferred examples thereof are now explained and set forth in detail.
The term xe2x80x9clowerxe2x80x9d means, unless otherwise indicated, a group having 1 to 6 carbon atoms.
Preferable examples of the xe2x80x9calkyl groupsxe2x80x9d and an alkyl moiety of the xe2x80x9calkoxy groupxe2x80x9d include a straight or branched chain aliphatic hydrocarbon residue, for example, a lower alkyl group such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl, neopentyl and hexyl.
Preferable examples of the xe2x80x9calkenyl groupsxe2x80x9d include a straight or branched chain aliphatic hydrocarbon residue having one double-bond, for example, a lower alkenyl group such as vinyl, propenyl (e.g., allyl group), butenyl, methylpropenyl, pentenyl and hexenyl.
Preferable examples of the xe2x80x9caryl groupsxe2x80x9d include phenyl, tolyl, xylyl, cumenyl, mesityl and naphthyl.
Preferable protective groups in the xe2x80x9cprotected hydroxy groupsxe2x80x9d and the xe2x80x9cprotected aminoxe2x80x9d are 1-(lower alkylthio)-(lower) alkyl group such as a lower alkylthiomethyl group (e.g., methylthiomethyl, ethylthiomethyl, propylthiomethyl, isopropylthiomethyl, butylthiomethyl, isobutylthiomethyl, hexylthiomethyl, etc.), more preferably C1-C4 alkylthiomethyl group, most preferably methylthiomethyl group;
trisubstituted silyl group such as a tri(lower)alkylsilyl (e.g., trimethylsilyl, triethylsilyl, tributylsilyl, tert-butyldimethylsilyl, tri-tert-butylsilyl, etc.) or lower alkyl-diarylsilyl (e.g., methyldiphenylsilyl, ethyldiphenylsilyl, propyldiphenylsilyl, tert-butyldiphenyl-silyl, etc.), more preferably tri(C1-C4)alkylsilyl group and C1-C4 alkyldiphenylsilyl group, most preferably tert-butyldimethylsilyl group and tert-butyldiphenylsilyl group; and an acyl group such as an aliphatic, aromatic acyl group or an aliphatic acyl group substituted by an aromatic group, which are derived from a carboxylic acid, sulfonic acid or carbamic acid.
Examples of the aliphatic acyl groups include a lower alkanoyl group optionally having one or more suitable substituents such as carboxy, e.g., formyl, acetyl, propionyl, butyryl, isobutyryl, valeryl, isovaleryl, pivaloyl, hexanoyl, carboxyacetyl, carboxypropionyl, carboxybutyryl, carboxyhexanoyl, etc.;
a cyclo(lower)alkoxy(lower)alkanoyl group optionally having one or more suitable substituents such as lower alkyl, e.g., cyclopropyloxyacetyl, cyclobutyloxypropionyl, cycloheptyloxybutyryl, menthyloxyacetyl, menthyloxypropionyl, menthyloxybutyryl, menthyloxypentanoyl, menthyloxyhexanoyl, etc.; a camphorsulfonyl group; or a lower alkylcarbamoyl group having one or more suitable substituents such as carboxy or protected carboxy, for example, carboxy(lower)alkylcarbamoyl group (e.g., carboxymethylcarbamoyl, carboxyethylcarbamoyl, carboxypropylcarbamoyl, carboxybutylcarbamoyl, carboxypentylcarbamoyl, carboxyhexylcarbamoyl, etc.), tri-(lower)alkylsilyl (lower) alkoxycarbonyl (lower) alkylcarbamoyl group (e.g., trimethylsilylmethoxycarbonylethylcarbamoyl, trimethylsilylethoxycarbonylpropylcarbamoyl, triethylsilylethoxycarbonylpropylcarbamoyl, tert-butyldimethylsilylethoxycarbonylpropylcarbamoyl, tri-methylsilylpropoxycarbonylbutylcarbamoyl, etc.) and so on.
Examples of the aromatic acyl groups include an aroyl group optionally having one or more suitable substituents such as nitro, e.g., benzoyl, toluoyl, xyloyl, naphthoyl, nitrobenzoyl, dinitrobenzoyl, nitronaphthoyl, etc.; and
an arenesulfonyl group optionally having one or more suitable substituents such as halogen, e.g., benzenesulfonyl, toluenesulfonyl, xylenesulfonyl, naphthalenesulfonyl, fluorobenzenesulfonyl, chlorobenzenesulfonyl, bromobenzenesulfonyl, iodobenzenesulfonyl, etc.
Examples of the aliphatic acyl groups substituted by an aromatic group include ar(lower)alkanoyl group optionally having one or more suitable substituents such as lower alkoxy or trihalo(lower)alkyl, e.g., phenylacetyl, phenylpropionyl, phenylbutyryl, 2-trifluoromethyl-2-methoxy-2-phenylacetyl, 2-ethyl-2-trifluoromethyl-2-phenylacetyl, 2-trifluoromethyl-2-propoxy-2-phenylacetyl, etc.
More preferable acyl groups among the aforesaid acyl groups are C1-C4 alkanoyl group optionally having carboxy, cyclo(C5-C6)alkoxy(C1-C4)alkanoyl group having two (C1-C4) alkyls at the cycloalkyl moiety, camphorsulfonyl group, carboxy-(C1-C4)alkylcarbamoyl group, tri (C1-C4) alkylsilyl (C1-C4) alkoxycarbonyl (C1-C4)-alkylcarbamoyl group, benzoyl group optionally having one or two nitro groups, benzenesulfonyl group having halogen, or phenyl (C1-C4) alkanoyl group having C1-C4 alkoxy and trihalo(C1-C4)alkyl group. Among these, the most preferable ones are acetyl, carboxypropionyl, menthyloxyacetyl, camphorsulfonyl, benzoyl, nitrobenzoyl, dinitrobenzoyl, iodobenzenesulfonyl and 2-trifluoromethyl-2-methoxy-2-phenylacetyl.
Preferable examples of the xe2x80x9c5- or 6-membered nitrogen, sulfur and/or oxygen containing heterocyclic ringxe2x80x9d include a pyrrolyl group and a tetrahydrofuryl group.
xe2x80x9cA heteroaryl which may be substituted by suitable substituentsxe2x80x9d moiety of the xe2x80x9cheteroaryloxy which may be substituted by suitable substituentsxe2x80x9d may be the ones exemplified for R1 of the compound of the formula of EP-A-532,088, with preference given to 1-hydroxyethylindol -5-yl, the disclosure of which is incorporated herein by reference.
The compounds (I) and its pharmaceutically acceptable salt for use in accordance with this invention are well known to have excellent immunosuppressive activity, antimicrobial activity and other pharmacological activities and, as such, be of value for the treatment or prevention of rejection reactions by transplantation of organs or tissues, graft-vs-host diseases, autoimmune diseases, and infectious diseases [EP-A-0184162, EP-A-0323042, EP-A-423714, EP-A-427680, EP-A-465426, EP-A-480623, EP-A-532088, EP-A-532089, EP-A-569337, EP-A-626385, WO89/05303, WO93/05058, WO96/31514, WO91/13889, WO91/19495, WO93/5059, etc.], the disclosures of which are incorporated herein by reference.
Particularly, the compounds which are designated as FR900506 (=FK506), FR900520 (ascomycin), FR900523, and FR900525 are products produced by microorganisms of the genus Streptomyces, such as Streptomyces tsukubaensis No. 9993 [deposited with National Institute of Bioscience and Human Technology Agency of Industrial Science and Technology (formerly Fermentation Research Institute Agency of Industrial Science and Technology ), at 1-3, Higashi 1-chome, Tsukuba-shi, Ibaraki, Japan, date of deposit Oct. 5, 1984, accession number FERM BP-927] or Streptomyces hygroscopicus subsp. yakushimaensis No. 7238 [deposited with National Institute of Bioscience and Human Technology Agency of Industrial Science and Technology (formerly Fermentation Research Institute Agency of Industrial Science and Technology ), at 1-3, Higashi 1-chome, Tsukuba-shi, Ibaraki, Japan, date of deposit Jan. 12, 1985, accession number FERM BP-928][(EP-A-0184162]. The FK506 (general name: tacrolimus) of the following chemical formula, in particular, is a representative compound. 
Chemical name: 17-allyl-1,14-dihydroxy-12-[2-(4-hydroxy-3-methoxycyclohexyl)-1-methylvinyl]-23,25-dimethoxy-13,19,21,27-tetramethyl-11,28-dioxa-4-azatricyclo[22.3.1.04,9] octacos-18-ene-2,3,10,16-tetraone
The preferable macrolides (I) is tacrolimus, ascomycin or its derivatives such as 33-epi-chloro-33-desoxyascomycin, which is disclosed in EP 427,680, example 66a. Other preferable compounds (I) are, for example, the compound of example 6d in EP569337, and the compound of example 8, EP626385.
The compounds shown in EP-0184162, EP323042, EP424714, EP427680, EP465426, EP474126, EP480623, EP484936, EP532088, EP532089, EP569337, EP626385, WO89/05303, WO93/05058, WO96/31514, WO91/13889, WO91/19495, WO93/5059, WO96/31514 and so on, are also exemplified as the preferable examples of the macrolides (I), the disclosures of which are incorporated herein by reference.
Further example of the immunosuppressant is cyclosporins, such as cyclosporin A, B, D, etc, which are shown in THE MERCK INDEX (12th edition), No. 2821, the disclosure of which is incorporated herein by reference.
Still further example of the immunosuppressant is an another type of macrolide which is called as rapamycin [THE MERCK INDEX (12th edition), No. 8288] and its derivatives. Preferable example of the derivatives is an O-substituted derivative in which the hydroxy in position 40 of formula A illustrated at page 1 of WO 95/16691, incorporated herein by reference, is replaced by xe2x80x94OR1 in which R1 is hydroxyalkyl, hydroalkoxyalkyl, acylaminoalkyl and aminoalkyl; for example 40-O-(2-hydroxy)ethyl-rapamycin, 40-O-(3-hydroxy)propyl-rapamycin, 40-O-[2-(2-hydroxy)ethoxy]ethyl-rapamycin and 40-O-(2-acetaminoethyl)-rapamycin. These O-substituted derivatives may be produced by reacting rapamycin (or dihydro or deoxo-rapamycin) with an organic radical attached to a leaving group (for example RX where R is the organic radical which is desired as the O-substituent, such as an alkyl, allyl, or benzyl moiety, and X is a leaving group such as CCl3C(NH)O or CF3SO3) under suitable reaction conditions. The conditions may be acidic or neutral conditions, for example in the presence of an acid like trifluoromethanesulfonic acid, camphorsulfonic acid, p-toluenesulfonic acid or their respective pyridinium or substituted pyridinium salts when X is CCl3C(NH)O or in the presence of a base like pyridine, a substituted pyridine, diisopropylethylamine or pentamethylpiperidine when X is CF3SO3. The most preferable one is 40-O-(2-hydroxy)ethyl rapamycin, which is disclosed in WO94/09010, the disclosure of which is incorporated herein by reference.
The macrolides (I), and rapamycin and its derivatives, have a similar basic structure, i.e., tricyclic macrolide structure, and at least one of the biological properties (for example, immunological properties).
The preferable immunosuppressant in the present invention is the one having the inhibitory activity on TNF-xcex1 and/or IFN-xcex3 production.
The immunosuppressant may be in a form of its salt, which includes conventional non-toxic and pharmaceutically acceptable salt such as the salt with inorganic or organic bases, specifically, an alkali metal salt such as sodium salt and potassium salt, an alkali earth metal salt such as calcium salt and magnesium salt, an ammonium salt and an amine salt such as triethylamine salt and N-benzyl-N-methylamine salt.
With respect to the immunosuppressant such as the macrolides (I), it is to be understood that there may be conformers and one or more stereoisomers such as optical and geometrical isomers due to asymmetric carbon atom(s) or double bond(s), and such conformers and isomers are also included within the scope of the present invention.
The immunosuppressant, particularly the macrolides (I) or its pharmaceutically acceptable salt, can be in the form of a solvate, which is included within the scope of the present invention. The solvate preferably include a hydrate and an ethanolate.
And further, the immunosuppressant can be in the form of pro-drugs, suitable derivatives, and so on.
The preferred examples of the immunosuppressant are the macrolides (I), wherein each of adjacent pairs of R3 and R4 or R5 and R6 independently form another bond formed between the carbon atoms to which they are attached;
each of R8 and R23 is independently a hydrogen atom;
R9 is a hydroxy group;
R10 is a methyl group, an ethyl group, a propyl group or an allyl group;
X is (a hydrogen atom and a hydrogen atom) or an oxo group;
Y is an oxo group;
each of R14, R15, R16, R17, R18, R19, and R22 is a methyl group;
R24 is a 3-R20-4-R21-cyclohexyl group,
in which R20 is hydroxy, an alkoxy group, an oxo group, or a xe2x80x94OCH2OCH2CH2OCH3 group, and
R21 is hydroxy, xe2x80x94OCN, an alkoxy group, a heteroaryloxy which may be substituted by suitable substituents, a xe2x80x94OCH2OCH2CH2OCH3 group, a protected hydroxy group, chloro, bromo, iodo, aminooxalyloxy, an azido group, p-tolyloxythiocarbonyloxy, or R25R26CHCOOxe2x80x94,
in which R25 is optionally protected hydroxy or protected amino, and
xe2x80x83R26 is hydrogen or methyl, or
R20 and R21 together form an oxygen atom in an epoxide ring; and
n is an integer of 1 or 2.
Tacrolimus is the most preferable compound belonging to the immunosuppressant. Other preferable compounds are listed herein below.
17-Ethyl-1,14-dihydroxy-12- [2-(4-hydroxy-3-methoxycyclohexyl)-1-methylvinyl]-23,25-dimethoxy- 13,19,21,27-tetramethyl-11,28-dioxa-4-azatricyclo[22.3.1.0 4,9]octacos-18-ene-2, 3,10,16-tetraone (=ascomycin): 33-epi-chloro-33-desoxyascomycin (EP-A-427680, example 66a): and 40-O-(2-hydroxy) ethyl rapamycin (WO94/0910)
xe2x80x9cMMP-production inhibitorxe2x80x9d in the present invention is used to mean the one having inhibitory or reducing activity on the production of MMPs. Preferable MMPs is gelatinase and collagenase. Most preferable xe2x80x9cMMP-production inhibitorxe2x80x9d is collagenase-production inhibitor.
xe2x80x9cMMP-mediated diseasesxe2x80x9d in the present invention is used to mean various diseases and pathological conditions caused by MMPs involving abnormal connective tissue and basement membrane matrix metabolism.
Particularly, preferable MMP-mediated diseases are the diseases or conditions caused by gelatinase and/or collagenase, and/or inflammatory diseases concerned with gelatinase; such as arthritis (e.g., osteoarthritis, rheumatoid arthritis, etc.), cerebral diseases (e.g., stroke, etc.), tissue ulceration (e.g., corneal, epidermal and gastric ulceration, etc.), abnormal wound healing, periodontal diseases, bone diseases (e.g., Paget""s diseases, osteoporosis, etc.), tumor growth, tumor metastasis or invasion, HIV-infection, decubitus, decubitis ulcer, restenosis, epidermolysis bullosa, sepsis, septic shock, neoplasm, psoriasis, neovascularization, multiple sclerosis, and soon. More preferable xe2x80x9cMMP-mediated diseasesxe2x80x9d of the present invention is cartilage degradation and/or connective tissue degradation, and rheumatoid arthritis which accompanies such degradation.
The immunosuppressant used in the present invention may be administered as pure compounds or mixtures of compounds or preferably, in a pharmaceutical vehicle or carrier.
The pharmaceutical compositions of this invention can be used in the form of a pharmaceutical preparation, for example, in solid, semisolid or liquid form, which contains the immunosuppressant, as an active ingredient, in admixture with an organic or inorganic carrier or excipient suitable for pharmaceutical use. The active ingredient may be compounded, for example, with the usual non-toxic, pharmaceutically acceptable, carriers for solutions (saline, for example), emulsion, suspensions (olive oil, for example), ointment, aerosol sprays, lotion, cream, gel, skin plasters, patches and any other form suitable for use. The carriers which can be used are water, glucose, lactose, gum acacia, gelatin, mannitol, starch paste, magnesium trisilicate, talc, cornstarch, keratin, colloidal silica, potato starch, urea and other carriers suitable for use in manufacturing preparations, in solid, semisolid, or liquid form, and in addition auxiliary, stabilizing, thickening and coloring agents and perfumes may be used. The active object compound is included in the pharmaceutical composition in an effective amount sufficient to produce the desired effect upon the process or condition of the disease.
Mammals which may be treated using the method of the present invention include livestock mammals such as cows, horses, etc., domestic animals such as dogs, cats, rats, etc. and humans.
While the dosage of therapeutically effective amount of the immunosuppressant varies from and also depends upon the age and condition of each individual patient to be treated, a daily dose of about 0.0001-1000 mg, preferably 0.001-500 mg and more preferably 0.01-100 mg. of the active ingredient is generally given for treating diseases, and an average single dose of about 0.001-0.01 mg, 0.2-0.5 mg, 1 mg, 5 mg, 10 mg, 50 mg, 100 mg, 250 mg and 500 mg is generally administered. Daily doses for chronic administration in humans will be in the range of about 0.1-0.3 mg/kg/day.